Functional divergence of zebrafish keap1 paralogs revealed by CRISPR/Cas9-mediated gene editing: a specialized role...

The Keap1/Nrf2 signaling pathway is a master regulator of cellular defense against oxidative and electrophilic stress. In teleosts like zebrafish (Danio rerio), whole-genome duplication resulted in two keap1 paralogs, keap1a and keap1b, whose functional specificities remain incompletely understood. This study investigates the divergent roles of these paralogs by comparing the responses of established keap1a and novel keap1b knockout larvae to distinct chemical stressors. By comparing the responses of keap1b^dl40, keap1a^dl07, and nfe2l2a^dl703 (Nrf2a) larvae to these stressors, we uncovered a striking functional dichotomy. While loss of either paralog conferred resistance to H₂O₂-induced oxidative stress, keap1b^dl40 larvae, unlike their keap1a^dl07 counterparts, exhibited extreme sensitivity to the lethal effects of CuSO₄ exposure, with survival rates plummeting to ~ 25%. This heightened sensitivity to copper sulfate was associated with a blunted transcriptional response of inflammatory markers tnf-a and c3a, suggesting that Keap1b is critical for modulating the Nrf2a-mediated response to inflammatory stress in orchestrating a viable inflammatory response. This work clarifies the non-redundant, vital function of Keap1b in the response to heavy metal-induced stress and provides a valuable genetic resource (keap1b^dl40 null allele) for future studies.